The present invention relates to compositions and methods for determining the genotype associated with an increased or decreased susceptibility to manic-depressive illness. The invention also provides a means to determine an individual""s increased or decreased risk of developing manic-depressive illness.
Genome screening efforts by several groups, designed to identify regions linked to bipolar disorder, have revealed evidence for potential susceptibility loci on chromosome 18. Berrettini (1994) Proc Natl Acad Sci USA 91:5918-5921, reported evidence for a susceptibility locus in the pericentromeric region of the chromosome. In a subsequent study on an independent pedigree series, Stine (1995) Am. J. Hum. Genet. 57:1384-1394, found support for the prior hypothesis on 18p (Berrettini et al., Proc Natl Acad. Sci. USA, 91:5918-5921, 1994). In the same study, Stine (1995) supra, presented evidence for a possible additional linkage on 18q. Recently, Freimer (1996) Nature Genet. 12:436-441, proposed a predisposing locus close to the telomere of 18q in Costa Rican kindreds. These reports suggest that the regions potentially implicated in bipolar disorder encompass a very large portion of chromosome 18.
In addition to bipolar disorder, more than 25 other diseases have been localized to chromosome 18, approximately 80% of which still await the discovery of the underlying defective gene (Overhauser et al., Cytogenet Cell Gene, 71:106-117, 1995; Online Mendelian Inheritance in Man (OMIM) (TM). (Database on line. 1995; URL: http://www3.ncbi.nlm.nih.gov/omim/. cited Jan. 19, 1996)). Since this chromosome has a genetic length estimated to be 150 cM [Cooperative Human Linkage Center (CHLC), Science 265:2049-2054, 1994], which includes about 4.5% of the total length of the genome, it is expected to encode several thousand genes. Approximately 40 genes have been mapped to this chromosome [Overhauser et al., 1995; Genome Database (GDB), URL: http.//gdbwww.gdb.org/gdb/browser/docs/topq.html greater than [database online]. (1990-). Updated daily (cited Jan. 19, 1996]; OMIM, [Database on line. 1995; cited Jan. 19, 1996]. Between 1993 and 1995, only 14 genes have been added to the list of chromosome 18 genes (Geurts van Kessel et al., Cytogenet Cell Gene, 65:141-165, 1994; Overhauser et al., Cytogenet Cell Gene, 71:106-117, 1995). Therefore, a dense transcriptional map, which would be valuable in positional cloning of susceptibility genes, remains to be developed for chromosome 18.
In one aspect the present invention is directed to a method for determining a genotype associated with increased susceptibility to manic-depressive illness. The method comprises determining the genotype of an affected individual with at least one polymorphic marker localized within the chromosomal region defined by and including markers D18S843 and D18S869, and determining therefrom the genotype associated with increased susceptibility to manic-depressive disorder.
In preferred embodiments the polymorphic marker is amplified by primers which selectively hybridize, under stringent conditions, to the same nucleic acid sequences as primers of SEQ ID NO:1 and SEQ ID NO:2 (see Table 1, below, forward and reverse primers to amplify Clone 22). Typically the polymorphic marker is amplified by the polymerase chain reaction.
In other embodiments the method of further comprises determining the genotype of a tested individual wherein the genotype is determined with at least one polymorphic marker localized within the chromosomal region defined by and including markers D18S843 and D18S869. The genotype of the tested individual is compared to the genotype associated with increased susceptibility to manic-depressive illness and the increased or decreased risk of the tested individual developing manic-depressive illness is
determined therefrom. Generally, the polymorphic marker of the tested individual is amplified by primers which selectively hybridize, under stringent conditions, to the same nucleic acid sequences as primers of SEQ ID NO:1 and SEQ ID NO:2.
In another aspect, the present invention is directed to a nucleic acid composition comprising oligonucleotide primers which selectively hybridize, under stringent conditions, to the same nucleic acid sequence as primers of SEQ ID NO:1 and SEQ ID NO:2. In an additional aspect the present invention is directed to a nucleic acid of less than 10 kB and comprising a polymorphic marker amplified by oligonucleotide primers of SEQ ID NO:1 and SEQ ID NO:2.
In yet another aspect, the present invention is directed to a method for determining an increased susceptibility to manic-depressive illness in an individual, comprising determining the genotype of the individual with oligonucleotide primers. The oligonucleotide primers amplify a polymorphic site as primers of SEQ ID NO:1 and SEQ ID NO:2. This polymorphic marker can be found in at least two forms, designated as xe2x80x9callele 1xe2x80x9d of clone 22 (SEQ ID NO:14) or xe2x80x9callele 2xe2x80x9d of clone 22 (SEQ ID NO:15). The presence of allele 2 of the polymorphic marker indicates an increased susceptibility to manic-depressive illness.
The invention further provides for a isolated nucleic acid encoding an IMP.18p myo-inositol monophosphatase, the protein defined as having a calculated molecular weight of between about 22 to 34 kDa, and where the protein""s activity includes hydrolysis of myo-inositol 1-phosphate to generate inositol and inorganic phosphate; and where the protein specifically binds to an antibody raised against an IMP.18p myo-inositol monophosphatase protein, or immunogenic fragment thereof, consisting of SEQ ID NO:17; or, having at least 60% amino acid sequence identity to an IMP.18p myo-inositol monophosphatase protein consisting of SEQ ID NO:17, as measured using a sequence comparison algorithm. In one embodiment, the nucleic acid encodes a IMP.18p myo-inositol monophosphatase having a calculated molecular weight of about 28 to 29 kDa. In other embodiments, the isolated nucleic acid: encodes a protein which has at least 80% amino acid sequence identity to the IMP. 18p myo-inositol monophosphatase protein of SEQ ID NO:17, as measured using a sequence comparison algorithm; encodes a protein having the sequence set forth in SEQ ID NO:17; specifically hybridizes to SEQ ID NO:16 under stringent conditions; or, encodes an IMP.18p myo-inositol monophosphatase protein which specifically binds to an antibody directed against a protein having a sequence as set forth in SEQ ID NO:17.
In further embodiments, the invention also provides for a polynucleotide or fragment thereof comprising a purified antisense nucleotide capable of hybridizing to and having a nucleic acid sequence complementary to at least a portion of an IMP.18p myo-inositol monophosphatase polynucleotide. The invention also provides for an expression vector comprising a nucleic acid encoding an IMP.18p myo-inositol monophosphatase or its antisense sequence. Further embodiments provide for a cell comprising an exogenous nucleic acid sequence encoding an IMP.18p myo-inositol monophosphatase protein. Another embodiment provides for an organism into which an exogenous nucleic acid sequence which specifically hybridizes under stringent conditions to SEQ ID NO:16 or which comprises a nucleic acid encoding an IMP.18p myo-inositol monophosphatase or fragment thereof, has been introduced, and the organism expresses the exogenous nucleic acid as an IMP.18p myo-inositol monophosphatase protein, or fragment thereof. In one embodiment, the organism""s exogenous nucleic acid sequence is translated into an IMP.18p myo-inositol monophosphatase protein which is expressed externally from the organism.
The invention also provides for an isolated IMP.18p myo-inositol monophosphatase protein having a calculated molecular weight of about 22 to 34 kDa; where the protein""s activity includes hydrolysis of myo-inositol 1-phosphate to generate inositol and inorganic phosphate; and specifically binds to an antibody raised against a myo-inositol monophosphatase protein, or immunogenic fragment thereof, consisting of SEQ ID NO:17, or has at least 60% amino acid sequence identity to a myo-inositol monophosphatase protein consisting of SEQ ID NO:17, as measured using a sequence comparison algorithm. In one embodiment, the isolated IMP.18p myo-inositol monophosphatase protein can also be found in humans. In further embodiments, the isolated IMP.18p myo-inositol monophosphatase protein has a calculated molecular weight of about 28 to 29 kDa; or, has a sequence as set forth in SEQ ID NO:17.
The invention further provides for an isolated antibody which is specifically immunoreactive under immunologically reactive conditions to an IMP.18p myo-inositol monophosphatase protein having the sequence as set forth in SEQ ID NO:17. In another embodiment, the isolated antibody is specifically immunoreactive under immunologically reactive conditions to an IMP.18p myo-inositol monophosphatase protein encoded by a IMP.18p myo-inositol monophosphatase nucleic acid of the invention.
Also provided for in the invention is a pharmaceutical composition comprising an acceptable carrier and an IMP.18p myo-inositol monophosphatase protein; an anti-IMP.18p myo-inositol monophosphatase antibody or binding fragment thereof; or a polynucleotide encoding an IMP.18p myo-inositol monophosphatase protein.
The invention also provides for a method for quantifying the amount of a myo-inositol monophosphatase in a mammal, comprising: obtaining a cell or tissue sample from the mammal; and, determining the amount of an IMP.18p myo-inositol monophosphatase gene product in the cell or tissue.
Another embodiment provides for a method for detecting the presence of a polynucleotide sequence encoding at least a portion of an IMP.18p myo-inositol monophosphatase in a biological sample, comprising the steps of providing a biological sample suspected of containing a IMP.18p myo-inositol monophosphatase-encoding nucleic acid and a probe capable of hybridizing to at least a portion of an IMP.18p myo-inositol monophosphatase nucleotide sequence, or a fragment thereof, from a biological sample; then combining the nucleic acid-containing biological sample with the probe under conditions such that a hybridization complex is formed between the nucleic acid and the probe; and detecting the hybridization complex. In one embodiment the nucleic acid in the biological sample is ribonucleic acid. In another embodiment, the detected hybridization complex correlates with expression of an IMP.18p myo-inositol monophosphatase in the biological sample.
The invention also provides for a method of determining whether a test compound is a modulator of an IMP.18p myo-inositol monophosphatase activity, the method comprising the steps of: providing a composition comprising an IMP.18p myo-inositol monophosphatase protein; contacting the monophosphatase with the test compound; and measuring the activity of the monophosphatase, wherein a change in monophosphatase activity in the presence of the test compound is an indicator of whether the test compound modulates monophosphatase activity. In one embodiment, the composition comprises monophosphatase is encoded a an IMP.18p myo-inositol monophosphatase polypeptide of the invention. In further embodiments, the composition comprises a cell or an organism.